药学学报, 2019, 54(7): 1200-1206
引用本文:
赵伟曼, 董智勇, 时宗芬, 鲁星月, 刘浩, 张配. 下调miR-205-5p增强鼻咽癌细胞HNE1/DDP对顺铂诱导凋亡的敏感性[J]. 药学学报, 2019, 54(7): 1200-1206.
ZHAO Wei-man, DONG Zhi-yong, SHI Zong-fen, LU Xing-yue, LIU Hao, ZHANG Pei. Down-regulation of miR-205-5p sensitizes HNE1/DDP to cisplatin induced apoptosis in vitro[J]. Acta Pharmaceutica Sinica, 2019, 54(7): 1200-1206.

下调miR-205-5p增强鼻咽癌细胞HNE1/DDP对顺铂诱导凋亡的敏感性
赵伟曼, 董智勇, 时宗芬, 鲁星月, 刘浩, 张配
蚌埠医学院药学院, 安徽 蚌埠 233030
摘要:
探讨下调miR-205-5p增强鼻咽癌耐药细胞HNE1/DDP对顺铂诱导凋亡的敏感性及其可能机制。qRT-PCR检测HNE1、HNE1/DDP细胞中miR-205-5p的表达差异及HNE1/DDP细胞转染miR-205-5p抑制剂后miR-205-5p的变化;MTT法检测顺铂(DDP)和miR-205-5p抑制剂对HNE1/DDP或HNE1细胞的增殖抑制作用;PI单染流式技术检测细胞凋亡情况;DAPI染色观察细胞核的形态;Western blot检测Bax、Bak、Mcl-1、Bcl-2蛋白的表达水平。结果显示,相比于HNE1细胞,HNE1/DDP高表达miR-205-5p,转染miR-205-5p抑制剂后其表达明显下降。下调miR-205-5p可显著提高HNE1/DDP细胞对顺铂的敏感性(P<0.05)。PI单染结果显示,转染miR-205-5p抑制剂可增强HNE1/DDP细胞对顺铂的诱导凋亡作用。miR-205-5p抑制剂联合DDP(8 μmol·L-1)作用HNE1/DDP细胞24 h的凋亡率为(28.93 ±2.50)%,相比单用miR-205-5p抑制剂(9.83 ±1.31)%或DDP的凋亡率(10.83 ±1.70)%显著升高(P<0.05)。联合作用组细胞表现为细胞核明显皱缩浓集呈碎片状。miR-205-5p抑制剂与DDP合用能上调Bax的表达和下调Bcl-2的表达。提示下调miR-205-5p可增强HNE1/DDP细胞对DDP的敏感性,其作用机制可能是促凋亡蛋白Bax的升高和抗凋亡蛋白Bcl-2的下降。
关键词:    鼻咽癌细胞      顺铂      miR-205-5p      耐药性      凋亡     
Down-regulation of miR-205-5p sensitizes HNE1/DDP to cisplatin induced apoptosis in vitro
ZHAO Wei-man, DONG Zhi-yong, SHI Zong-fen, LU Xing-yue, LIU Hao, ZHANG Pei
School of Pharmacy, Bengbu Medical College, Bengbu 233030, China
Abstract:
This study aims to investigate the effect of down-regulation of miR-205-5p by transfection of miR-205-5p inhibitor on the sensitivity of HNE1/DDP cells to cisplatin (DDP) induced apoptosis and explore the underlying mechanism. qRT-PCR was used to detect the expression of miR-205-5p in HNE1 or HNE1/DDP cells. The expression level of miR-205-5p was analyzed after transfecting HNE1/DDP cells with miR-205-5p inhibitor. MTT assay was used to evaluate the inhibitory effect of DDP alone or in combination with miR-205-5p inhibitor on the proliferation of HNE1/DDP or HNE1 cells. Apoptosis of cells treated with miR-205-5p inhibitor alone or in combination with DDP (8 μmol·L-1) was assessed using flow cytometry with PI staining, with the nucleus was counterstained with DAPI staining. The expression of Bax, Bak, Mcl-1, or Bcl-2 was analyzed by Western blot. HNE1/DDP cells showed a high level of expression of miR-205-5p, and the expression of miR-205-5p was significantly decreased by transfection of miR-205-5p inhibitor. Down-regulation of miR-205-5p significantly increased the sensitivity of HNE1/DDP cells to DDP (P<0.05). Transfection of miR-205-5p inhibitor enhanced the sensitivity of HNE1/DDP cells to DDP induced apoptosis. Treatment of HNE1/DDP cells with miR-205-5p inhibitor combined with DDP (8 μmol·L-1) for 24 h resulted in an apoptotic rate of 28.93% ±2.50%, significantly higher than that treated with miR-205-5p inhibitor (9.83% ±1.31%) or DDP alone (10.83% ±1.70%) (P<0.05). DAPI staining showed that HNE1/DDP cell nucleus became significantly condensed and fragmented in miR-205-5p inhibitor combined with DDP group. The combined group up-regulated the expression of Bax and down-regulated the expression of Bcl-2 in HNE1/DDP cells. Therefore, down-regulation of miR-205-5p can enhance the sensitivity of HNE1/DDP cells to cisplatin induced apoptosis, and the mechanism may involve up-regulation of Bax and down-regulation of Bcl-2 expression.
Key words:    nasopharyngeal carcinoma cell    cisplatin    miR-205-5p    drug resistance    apoptosis   
收稿日期: 2018-12-24
DOI: 10.16438/j.0513-4870.2018-1142
基金项目: 国家自然科学基金资助项目(81603155);蚌埠医学院自然科学基金资助项目(BYKY1763).
通讯作者: 张配,Tel:86-552-3175452,E-mail:zhangpei880204@126.com
Email: zhangpei880204@126.com
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